This invention relates to methods of treatment using MAO-A or MAO-B inhibitors such as L-deprenyl.
L-Deprenyl, also known as selegiline or Eldepryl, is a selective inhibitor of mitochondrial monoamine oxidase type B (MAO-B). It belongs to a class of enzyme-activated irreversible inhibitors also described as xe2x80x9csuicidexe2x80x9d inhibitors, because the compound acts as a substrate for monoamine oxidase, the action of which on the compound results in irreversible inhibition. L-Deprenyl forms a monovalent complex with monoamine oxidase as an initial, reversible step. Subsequent interaction of L-deprenyl with MAO leads to a reduction of the enzyme-bound flavine-adenine dinucleotide (FAD), and concomitant oxidation of the inhibitor. The oxidized inhibitor then reacts with FAD at the N-5-position in a covalent manner.
L-Deprenyl has been used clinically as an MAO-B inhibitor in combination with levo-dopa (L-dopa) to treat Parkinson""s disease. L-Dopa treatment alone is optimally effective only for the first few years of therapy. The anti-Parkinson""s disease action of L-deprenyl was based on the theory that MAO-B was the predominant form of MAO in the brain and that brain MAO rather than peripheral enzyme activity was to be selectively inactivated. Use of L-deprenyl in conjunction with L-dopa therapy enhances dopaminergic transmission. This permits a lowering of the dosage of L-dopa, which prolongs the effect of L-dopa and decreases adverse side effects of L-dopa.
L-Deprenyl has been reported to enhance catecholaminergic activity and diminish serotoninergic activity in the brain, by mechanisms unrelated to MAO-B inhibition. In rats it has been shown to reduce brain damage after exposure to transient hypoxia-ischemia, the proposed mechanism being either a prevention of the rise of H2O2 or an increase in enzymatic radical scavenging capacity, particularly by facilitating superoxide dismutase activity. Indeed, some of these additional mechanisms may contribute to L-deprenyl""s mode of action in Parkinson""s disease.
It is an object of the present invention to provide a method of increasing nitric oxide production.
It is a further object of the present invention to provide a method of treating diseases of brain and blood vessels related to a deficiency in nitric oxide production.
It is yet another object of the present invention to provide a method of protecting the vascular endothelium.
It is a further object of the present invention to provide a method of relaxing non-vascular smooth muscle.
It is another object of the present invention to provide a method of treating neuronal disorders.
It is a further object of the present invention to provide a method of treating cellular disorders of platelets, RBC, WBC, mast cells, macrophages, and glial cells.
These and other objects of the invention are provided by a method of treating a disorder of the vasculature, comprising administering to a subject suffering from such a disorder an effective amount of an MAO-A or MAO-B inhibitor. The disorder of the vasculature may be a disorder of the cerebral or peripheral vasculature. Specific vasculature disorders which can be treated include essential, renovascular, pulmonary, and ocular hypertension, myocardial infarction, and cerebrovascular stroke.
The disorder of the vasculature may be associated with a deficiency in NO production, with the MAO-A or MAO-B inhibitor exerting an endothelium-dependent effect on the vasculature. Alternatively, the disorder may be a disorder of the vasculature associated with a toxic effect of xcex2-amyloid on the vasculature, in which case the MAO-A or MAO-B inhibitor protects the endothelium of the vasculature from xcex2-amyloid. The disorder also may be one not associated with a deficiency in NO production, in which case the MAO-A or MAO-B inhibitor exerts an endothelium-independent effect on the vasculature.
The present invention also provides a method of treating a neuronal disorder other than Parkinson""s Disease or Alzheimer""s Disease, comprising administering to a subject suffering from such a disorder an effective amount of an MAO-A or MAO-B inhibitor. The disorder may be associated with a deficiency in NO production, in which case the MAO-A or MAO-B inhibitor stimulates production of NO. Alternatively, the neuronal disorder may be caused by a toxic effect of xcex2-amyloid on neurons.
Also provided according to the invention is a method of treating a disorder of the non-vascular smooth muscle, comprising administering to a subject suffering from such a disorder an effective amount of an MAO-A or MAO-B inhibitor. Disorders of the non-vascular smooth muscle that can be treated include airway obstruction or another respiratory disorder and a gastrointestinal motility disorder.
The present invention also provides a method of treating a cellular disorder of platelets, RBC, WBC, mast cells, macrophages, or glial cells, comprising administering to a subject suffering from such a disorder an effective amount of an MAO-A or MAO-B inhibitor. In a preferred embodiment, the MAO-A or MAO-B inhibitor acts as an anti-platelet agent or an anti-inflammatory agent.
In preferred embodiments, the MAO-A or MAO-B inhibitor is selected from the group consisting of L-deprenyl, clorgyline, pargyline, N-(2-aminoethyl)-4-chlorobenzamide hydrochloride, N-(2-aminoethyl)-5(3-fluorophenyl)-4-thiazolecarboxamide hydrochloride, and derivatives thereof. A dose of 1-100 mg/day, preferably 1-10 mg/day, of the MAO-A or MAO-B inhibitor is used in accordance with the present invention.
Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.
It has been discovered, surprisingly, that MAO-A or MAO-B inhibitors such as L-deprenyl display many modes of action which are totally unrelated to their mode of action as selective inhibitors of MAO-A and/or MAO-B. For example, it has been found that MAO-A or MAO-B inhibitors such as L-deprenyl exert effects on both cerebral and peripheral vasculature, some of which are mediated by nitric oxide (NO) and others of which are NO-independent. More particularly, MAO-A or MAO-B inhibitors such as L-deprenyl have been found to stimulate NO production rapidly and stereospecifically when administered in vitro or in vivo to peripheral or cerebral blood vessels. They also have been found to blunt the vasoconstriction caused by a number of vasoconstrictors.
For example, L-deprenyl at low doses (xe2x89xa610 xcexcM) causes a rapid NO-mediated endothelium-dependent vasodilation. At higher doses L-deprenyl produces a slow progressive NO- and endothelium-independent direct relaxation of vascular smooth muscle. The NO-mediated, endothelial-dependent effects of L-deprenyl and other MAO-A or MAO-B inhibitors on the cerebral and peripheral vasculature makes them useful in treating a variety of disorders, including essential, renovascular and pulmonary hypertension, glaucoma (by reduction of intraocular pressure), macular degeneration, and erectile impotence all of which result from a significant reduction of endothelium-dependent relaxation. The NO-mediated, endothelial-dependent effects also are useful in preserving organs for transplantation. The blood-brain barrier is composed of endothelial cells, and by protecting the endothelium L-deprenyl and other MAO-A or MAO-B inhibitors protect the integrity of the blood-brain barrier. They also are useful in cases of myocardial infarction and cerebrovascular stroke which result from an alteration of endothelial function. The endothelium-independent direct relaxation of vascular smooth muscle by MAO-A or MAO-B inhibitors such as L-deprenyl can be a useful adjunct in treatment of these disorders.
MAO-A or MAO-B inhibitors such as L-deprenyl also have been discovered to exert a potent relaxant effect on non-vascular smooth muscle, which is mediated both by guanylate cyclase and cyclic GMP-independent mechanisms. This action makes them useful for treating disorders associated with relaxation of smooth muscle, such as airway obstruction and other respiratory disorders, gastrointestinal motility disorders, hemorrhoids, sphincter and smooth muscle spasm in the gastrointestinal tract, and bladder dysfunction. They may be used to counteract premature labor and to relax the birth canal during delivery. They also are useful in relaxing the urinary tract for the passage of kidney stones, and may be used to alleviate smooth muscle contraction and spasm, thus facilitating diagnostic procedures such as endoscopy, bronchoscopy, laparoscopy, cystoscopy and catheterization.
It additionally has been found that L-deprenyl and other MAO-A or MAO-B inhibitors exert NO-mediated effects on the nervous system that are unrelated to their action on MAO-A or MAO-B, leading to their use in treatment of neuronal disorders other than Parkinson""s Disease. Neuronal disorders caused by a deficiency in NO production include age-related neurodegenerative diseases, as well as other memory disorders. Exemplary of disorders that can be treated effectively with L-deprenyl and other MAO-A or MAO-B inhibitors are Alzheimer""s disease, Down""s syndrome, amyotrophic lateral sclerosis (ALS), Huntington""s disease, AIDs dementia, brain trauma, and learning and movement disorders. The compounds also can be used in neuron protection.
Finally, it has been discovered that MAO-A or MAO-B inhibitors such as L-deprenyl affect a diverse group of cells other than endothelial, non-vascular smooth muscle and neuronal cells. These include both NO-mediated and NO-independent effects on platelets, RBC, WBC, mast cells, macrophages, and glial cells. Most notable in this context are their use as anti-platelet agents or as anti-inflammatory agents. Examples of treatable disorders include asthma and thrombosis.
Thus, MAO-A or MAO-B inhibitors such as L-deprenyl can be used to effect both NO-mediated and NO-independent actions on the cerebral and peripheral vasculature, on non-vascular smooth muscle, and on a diverse group of other cells. They also are useful in treating neuronal tissues, including brain tissue, which suffer from a deficiency in NO production.
Activity has been demonstrated for a wide variety of MAO-A and MAO-B inhibitors, including L-deprenyl, clorgyline, pargyline, RO-16-6491 (N-(2-aminoethyl)-4-chlorobenzamide hydrochloride), and RO-41-1049 (N-(2-aminoethyl)-5-(3-fluorophenyl)-4-thiazolecarboxamide hydrochloride). Each of these compounds has been shown to have the ability to inhibit contraction, to dilate blood vessels, to inhibit xcex2-amyloid and to stimulate NO production in the brain. Derivatives of these compounds may be used, as well as other MAO-A and MAO-B inhibitors and derivatives thereof. Exemplary of compounds that are structurally related to L-deprenyl are N-propargylamine compounds, N-methyl-propargylamine and N-methyL-N-(2-pentyl)-propargylamine can be used in place of L-deprenyl.
The effect of these compounds on vasodilation has been confirmed in both peripheral and cerebral blood vessels. Low concentrations of the compounds produce relaxation of aortic rings with intact endothelium. This effect is not detectable in endothelium-free tissue and is greatly diminished in endothelium-intact tissue treated with the nitric oxide synthase (NOS) inhibitor L-Nitro-Arginine-Methyl-Ester, (L-NAME), which exhibits an enhanced vasoconstriction. Thus the vasodilatory effect of low concentrations of the compounds appears to be mediated by endothelial NOS.
Further evidence of the NO-mediated, endothelium-dependent vasodilatory effect of these compounds is demonstrated by the fact that pretreatment with freshly prepared hemoglobin blocks the vasodilatory effect of low concentrations of the compounds in rat aorta and bovine cerebral artery. By binding NO, hemoglobin prevents the vasodilatory action of NO. The endothelium-dependent effect of the compounds also is prevented by methylene blue (10xe2x88x924 M). Methylene blue is an inhibitor of the enzyme guanylate cyclase which catalyzes the formation of cyclic GMP, which in turn mediates the vasodilatory effect of NO in vascular smooth muscle.
NO or a labile NO-containing compound is considered to be the endothelium-derived relaxing factor (EDRF), which plays a vital role in the regulation of vascular tone. However, the endothelial-mediated vasodilation induced by MAO-A or MAO-B inhibitors such as L-deprenyl appears not to be mediated by any of the classical endothelial receptors. It is present even when different vasoconstrictors are used, e.g. phenylephrine, norephrine, 5-HT, the prostaglandin agonist U46619, or K+. The vasodilatory effect also appears not to be mediated by cholinergic receptors on endothelium since the antagonist atropine (10xe2x88x924 M) M) does not abolish the vasodilatory effect of L-deprenyl and other MAO-A or MAO-B inhibitors. Nor is the effect prostaglandin-mediated, since the cyclooxygenase inhibitor indomethacin (10xe2x88x925 M) fails to antagonize L-deprenyl-mediated vasodilation. The phospholipase A2 inhibitor manolide (10-5 M) also has no detectable effect on L-deprenyl-mediated vasodilation.
The possibility of amphetamine metabolites mediating the action of L-deprenyl can be eliminated. The D-isomer is metabolized to D-amphetamine which has 10 times the amphetamine potency of the L-amphetamine derived from L-deprenyl, yet the D-isomer is of much lower potency than the L-isomer.
High concentrations of L-deprenyl and other MAO-A and MAO-B inhibitor, e.g.,  greater than 2.5xc3x9710xe2x88x925 M for L-deprenyl, cause a slowly developing relaxation in endothelium-denuded aorta. This effect is not reversed by L-NAME, demonstrating that high concentrations of L-deprenyl directly relax vascular smooth muscle through an NO-independent mechanism as opposed to the NO-mediated, endothelium-dependent relaxation caused by low concentrations of the drug.
Rat and bovine cerebral vessels show a vasodilatory effect similar to that shown by aortic rings. Cerebral arteries with intact endothelium show a rapid vasodilatory response to low doses of L-deprenyl and other MAO-A and MAO-B, e.g., xe2x89xa610 xcexcM for L-deprenyl, which is prevented by L-NAME. As in peripheral blood vessels, in endothelium-denuded cerebral arteries higher doses of L-deprenyl and other MAO-A and MAO-B inhibitors induce a slow progressive relaxation which is not reversed by L-NAME.
The direct vasodilatory effect of L-deprenyl and other MAO-A or MAO-B inhibitors on vascular smooth muscle has similarities to the activation of calcium-dependent potassium channels and is probably mediated through ion channels. Both the endothelium-dependent and independent vasodilatory effects of L-deprenyl are abolished in calcium-free medium. The constitutive form of the enzyme NO synthase present in endothelial cells and neurons requires calcium for activation. Thus, the generation of NO by L-deprenyl and other MAO-A or MAO-B inhibitors involves the activation of constitutive NO synthase.
Antagonism of vasoconstriction by L-deprenyl extends to antagonism of vasoconstriction caused by a number of vasoconstrictors, as demonstrated by its action on isolated segments of blood vessels maintained in tissue bath. Rat aortic rings with intact endothelium, when pretreated with L-deprenyl or other MAO-A or MAO-B inhibitors, exhibit a dose-dependent decrease in contraction in response to the vasoconstrictor PE. Contractions at lower doses of PE are more sensitive to the effect of the compounds. Surprisingly, D-deprenyl actually enhances vasoconstriction, demonstrating the superior ability and stereospecific action of L-deprenyl in antagonizing PE-induced contraction.
Pretreatment of the intact aortic rings with the NOS inhibitor L-NAME abolishes the inhibitory action of L-deprenyl and other MAO-A and MAO-B inhibitors on PE-induced contraction. The inactive isomer D-Nitro-Arginine-Methyl-Ester (D-NAME) is less effective in blocking the effect of L-deprenyl. L-Deprenyl displays a similar inhibition of vasoconstriction induced by serotonin in bovine mid-cerebral artery. Inhibition of vasoconstriction by 10xe2x88x924 M L-deprenyl also is observed in endothelium-denuded peripheral and cerebral arteries.
The vascular effects of L-deprenyl and other MAO-A or MAO-B inhibitors have been confirmed in vivo. Aortas removed from rats one hour following injection with L-deprenyl or another MAO-A or MAO-B inhibitors or saline show diminished vasoconstriction in response to the vasoconstrictor PE and enhanced vasodilatory response to the vasodilator acetylcholine. The enhanced response to acetylcholine is evident from an observed increased response to low concentrations of acetylcholine as compared to control rats treated with saline. The results indicate that the effect of L-deprenyl and other MAO-A or MAO-B inhibitors on the vasculature persist for a significant period of time.
The discovery of the previously unknown effects of L-deprenyl and other MAO-A or MAO-B inhibitors on the endothelium, particularly the low-dose NO-mediated effects, leads directly to a method of protecting the endothelium from xcex2-amyloid toxicity. The present inventor has demonstrated that xcex2-amyloid causes endothelial dysfunction. Damaged endothelium exhibits an enhanced response to vasoconstrictors and diminished sensitivity to vasodilators. This includes enhanced vasoconstriction with serotonin and diminished relaxation to the endothelium-dependent vasodilators acetylcholine and bradykinin. Amyloid-mediated vascular damage is postulated to be an early event relative to development of Alzheimer""s Disease. Administration of L-deprenyl and other MAO-A or MAO-B inhibitors can prevent the vascular damage caused by xcex2-amyloid, thereby delaying the progression of Alzheimer""s Disease. Administration of compounds according to the invention also is effective in combating amyloid angiopathy.
The ability of L-deprenyl and other MAO-A or MAO-B inhibitors to prevent vascular damage caused by xcex2-amyloid has been confirmed by studies with rat aorta with the xcex2-amyloid peptide primarily associated with cerebrovascular deposits in Alzheimer""s disease, and the major circulating form of amyloid. xcex2-amyloid produces a significant increase in vasoconstriction induced by PE in rat aorta, and a diminished relaxation response to acetylcholine. Both effects are antagonized by pretreatment with L-deprenyl and other MAO-A or MAO-B inhibitors. Moreover, the aortas from rats injected with L-deprenyl and other MAO-A or MAO-B inhibitors do not exhibit any of the typical features of xcex2-amyloid-mediated endothelial dysfunction.
L-Deprenyl and other MAO-A or MAO-B inhibitors also antagonize the effect of xcex2-amyloid in bovine mid-cerebral arteries. xcex2-amyloid produces significantly increased vasoconstriction in response to serotonin, which is completely blocked by pretreatment with L-deprenyl and other MAO-A or MAO-B inhibitors. L-Deprenyl and other MAO-A or MAO-B inhibitors also antagonize the diminished vasodilatory response of the cerebral artery induced by xcex2-amyloid.
It is postulated that the cytoprotective effect of L-deprenyl and other MAO-A or MAO-B inhibitors is mediated by increased NO production, but it may be due in part to oxygen free radical scavenging by the compounds. Regardless of the mechanism, the ability of the compounds to antagonize xcex2-amyloid-mediated endothelial dysfunction in peripheral and cerebral blood vessels provides a significant treatment for preventing the vascular damage caused by xcex2-amyloid, particularly in Alzheimer""s Disease.
In addition to its effects on vascular and non-vascular smooth muscle, including those involving xcex2-amyloid, NO mediates a wide range of physiological activities, particularly certain forms of learning and memory. NO has a role in the cellular basis of memory by facilitating long-term potentiation (LTP). During LTP induction in the CA1 region of the hippocampus, NO generated in the dendrites of pyramidal cells transmits retrograde signals from the postsynaptic to the presynaptic terminals. The endothelial isoform of NOS seems to be the major enzyme involved in the maintenance of LTP in the hippocampus. NO also is involved in neuron protection.
NO mediates some of the effects of glutamate/NMDA receptor pathway on neuronal functioning and synaptic plasticity. L-Deprenyl and other MAO-A or MAO-B inhibitors stimulate NO production in all brain regions examined as well as in the mid-cerebral artery. L-Deprenyl is a particularly potent stimulant of NO production, being more effective in rat brains than the endogenous NOS stimulant L-glutamate: in all brain regions except the hippocampus, L-deprenyl stimulates NO at a more than 100-fold lower concentration than L-glutamate. In bovine cerebellum both 10 and 100 mM L-deprenyl produce significantly more NO than 1 mM L-glutamate, and concentrations as low as 1 mM have a stimulatory effect on NO production. In each case, D-deprenyl has considerably less activity.
L-Deprenyl and other MAO-A and MAO-B inhibitors can be administered orally in capsule form. For example, a recommended regimen for administration in accordance with the present invention is 1-100 mg/day, preferably divided among two or more doses. A preferred dosage is 10-20 mg/day.